Lyme borreliosis, or Lyme disease, is the most commonly reported tick-borne disease in Europe and North America. The disease is caused by the arthropod-borne spirochete Borrelia burgdorferi sensu lato (B. burgdorferi s.l.) and is a multi-systemic infection that can involve multiple organs or tissues, resulting in skin, cardiac, neurological and musculoskeletal disorders. B. burgdorferi s.l. can be divided into 12 genospecies. These 12 genospecies occur in different geographic regions, and live in nature in enzootic cycles involving ticks of the Ixodes ricinus complex (also called Ixodes persulcatus complex) and a wide range of animal hosts (Table A). From these 12 genospecies at least three are pathogenic for humans: Borrelia burgdorferi sensu stricto (B. burgdorferi s.s.), B. afzelii and B. garinii. Two other genospecies, B. lusitaniae and B. valaisiana, have occasionally been detected in humans and their role in Lyme borreliosis are still uncertain.
TABLE AThe B. burgdorferi s.l. genospecies, their tick vectors andgeographic location.Principal tick vectorLocationThree pathogenic speciesBorrelia burgdorferi s.s.Ixodes scapularisNorth-eastern andnorth central USIxodes pacificusWestern USIxodes ricinusEuropeBorrelia gariniiIxodes ricinusEuropeIxodes persulcatusAsiaBorrelia afzeliiIxodes ricinusEuropeIxodes persulcatusAsiaNine minimallypathogenic or non-pathogenic speciesBorrelia andersoniiIxodes dentatusEastern USBorrelia bissettiiIxodes spinipalpisWestern USIxodes pacificusBorrelia valaisianaIxodes ricinusEurope and AsiaBorrelia lusitaniaeIxodes ricinusEuropeBorrelia spielmaniIxodes ricinusEuropeBorrelia japonicaIxodes ovatusJapanBorrelia tanukiiIxodes tanukiiJapanBorrelia turdaeIxodes turdusJapanBorrelia sinicaIxodes persulcatusChina
Lyme borreliosis was described as a new clinical entity in 1976. Allen C. Steere investigated a group of people with rashes and swollen joints in Old Lyme, Conn., and misdiagnosed Lyme borreliosis as juvenile rheumatoid arthritis (Steere et al., 1977). The disease has, however, been known in Europe under a variety of names since the 1880's. In the year 1883, Buchwald described a chronic skin change, which received the name acrodermatitis chronica atrophicans (ACA). In the year 1910 the Swedish physician Afzelius discovered a circularly moving red rash after an insect or tick bite, which received the name erythema migrans (EM). Neurological symptoms after a tick bite were described 1922 by the French researchers Garin and Bujadoux. In 1951 the Swedish clinician Hollstrom to successfully treated patients with EM with penicillin and in 1984 Borrelia cells were observed in skin biopsies taken from an EM lesion. It was suggested that EM in association with meningitis probably was the result of an infection by a tick-borne or an insect-borne bacterium. The causative agent of Lyme borreliosis was finally discovered 1982 by W. Burgdorfer and colleagues, who isolated a previously unidentified spirochetal bacterium from the hard tick Ixodes scapularis that was later named Borrelia burgdorferi (Burgdorfer et al., 1982).
Epidemiology of Lyme Borreliosis
Lyme borreliosis is the most common tick-borne zoonosis in Europe and North America. While in most countries it is not a notifiable disease, no exact data are available how many new cases there are per year. In the United States the causative agent is B. burgdorferi s.s. and Lyme borreliosis is localized in north-eastern, mid-Atlantic and upper north-central states. In 2002, a total of 23,763 cases of Lyme borreliosis were reported to CDC, yielding a national incidence of 8.2 cases per 100,000 inhabitants (Lyme Disease Surveillance in Morbidity Mortality Weekly Reports, 2004). B. afzelii and B. garinii are the main causative agents of Lyme borreliosis in Europe together with B. burgdorferi s.s. which contributes to a lesser extent dependent on the geographic location. In Europe the incidence of Lyme borreliosis differs between countries. The average incidence of Lyme borreliosis in Eastern Germany was 17.8 cases per 100,000 population in 2002 and increased by 31% to 23.3 cases in 2003, respectively. In Austria, the incidence of Lyme borreliosis is much higher with 130 cases per 100,000 inhabitants (Report of WHO workshop on Lyme borreliosis and Diagnosis and surveillance, Warsaw, Poland, 20-22 June, 1995).
In some risk groups, such as farmers, forestry workers, hikers, runners or vacationers, seroprevalence and disease incidence rates have increased, as in children under 15 and adults between 39 and 59, without gender preference. The prevalence of Lyme borreliosis varies considerably in different European countries with an overall increased prevalence from west to east. The high incidence of Lyme disease is linked with changes in the forest habitats as well as social factors. An environmental change such as forest fragmentation has led to a loss of rodent predators such as wolves and birds of prey which in turn has led to an increase in the mouse population and automatically to an increase in the tick population. More recently patchy reforestation has increased the numbers of deer and thus the numbers of ticks. Middle class suburban sprawl and the increasing use of these woodland areas for recreation such as camping and hiking has brought humans into greater contact with the larger number of infected ticks. All these parameters contribute to a wider distribution of Lyme borreliosis.
Etiological Agent of Lyme Borreliosis
B. burgdorferi s.l. belongs to the family Spirochaetaceae, which is subdivided into the medically important genera Treponema, Leptospira and Borrelia. B. burgdorferi s.l. is a spiral shaped (10-20 μm long and 0.2-0.5 μm wide), vigorously motile, gram negative bacterium that grows under microaerophilic conditions. The spirochetal cell wall consists of a cytoplasmic membrane surrounded by peptidoglycan and flagella and then by a loosely associated outer membrane. The borrelial genome is perhaps the most structurally complex among bacteria and consists of one linear chromosome and a varying number of both linear and circular plasmids (Xu et al., 1995).
In 1997 the genome of B. burgdorferi s.s. strain B31 was completely sequenced by TIGR (Fraser et al., 1997), and the following analysis showed that the 1.5 Mb genome sequence is encoded in a single chromosome plus a highly dynamic complement of 12 linear and 9 circular plasmids (Casjens et al., 2000). The genomic sequence of the B. garinii strain PBi was published in 2004 (Glockner et al., 2004). They were only able to assemble two plasmids completely, the remaining plasmids sequence are left as variable plasmid sequences, indicating the complexity and the high content of repetitive sequences. There is a 92% nucleotide sequence identity between the genomes from B. burgdorferi s.s. strain B31 and B. garinii strain PBi, the sequences of the plasmids are more divergent.
About 90% of the chromosomal genomes comprises coding regions and most of the encoded genes are homologous to genes of known function. They code for proteins involved in DNA replication, transcription and translation; the repair system and recombination; transport, nutrient uptake and energy metabolism; motility and chemotaxis and the regulation of gene to expression. Genes related with pathogenicity are primarily located on the extra-chromosomal genome. The genes coding for the synthesis of amino acids, fatty acids, cofactors and nucleotides are absent from the borrelial genome. Therefore Borrelia require a complex medium supplemented with serum, such as BSK-II medium, for growth in vitro.
Clinical Manifestations of Lyme Borreliosis
Lyme borreliosis generally occurs in stages, with remission and exacerbations with different clinical manifestation at each stage (Steere, 1989). Early infection stage 1 consists of localized infection of the skin, followed within days or weeks by stage 2, disseminated infection, and months to years later by stage 3, persistent infection. However, the infection is variable; some patients have only localized infections of the skin, while others display only later manifestations of the illness, such as arthritis. Different clinical syndromes of Lyme borreliosis are also caused by infection with diverse B. burgdorferi s.l. species. B. burgdorferi s.s. more often causes joint manifestations (arthritis) and heart problems, B. afzelii causes mainly dermal symptoms (EM and ACA), and B. garinii is mostly responsible for neuroborreliosis.
Localized infection—The most common symptom of stage 1 of an infection is erythema migrans, which occurs in 70-80% of infected people. This skin lesion is often followed by flu-like symptoms, such as myalgia, arthralgia, headache and fever. These non-specific symptoms occur in 50% of patients with erythema migrans.
Disseminated infection—During stage 2 the bacteria move into the blood stream from the site of infection and to more distant tissues and organs. Neurological, cardiovascular and arthritic symptoms that occur in this stage include meningitis, cranial neuropathy and intermittent inflammatory arthritis.
Persistent infection—Stage 3 of the infection is chronic and occurs from months to years after the tick bite. The most common symptom in North America is rheumatoid arthritis, caused by an infection with B. burgdorferi s.s. Persistent infection of the central nervous system with B. garinii causes more severe neurological symptoms during stage 3, and a persistent infection to of the skin with B. afzelii results in acrodermatitis chronica atrophicans.
Diagnosis and Treatment
The best parameter for diagnosis of early Lyme borreliosis is the characteristic expanding red skin lesion or EM. Unfortunately, 30 percent of patients do not develop this rash, and the usual symptoms of early disease, such as fatigue, fever, or headache, are too non-specific to be diagnostic. Lyme borreliosis in a symptomatic patient can be diagnosed using a variety of serological tests, and the most common diagnostic test is indirect ELISA testing for Lyme specific antibodies in serum.
Antimicrobial agents are the principle method of treatment of Lyme borreliosis infection. The antibiotic used depends on the stage of the disease, symptoms, and the patient's allergies to medication. The length of the antibiotics course also depends on the stage of the disease and severity of symptoms. Early Lyme borreliosis is typically treated with oral tetracyclines, such as doxycycline, and semi-synthetic penicillins, such as amoxicillin or penicillin V. Arthritic and neurological disorders are treated with high-dose intravenous penicillin G or ceftriaxone.
Prevention
The number of cases of Lyme borreliosis steadily increases, as a result of changed ecological conditions. In addition, the difficulty of prevention indicates that the incidence will continue to be a public health concern. Landscape modifications, protective clothing, tick checks or personals protection measures are helpful but not enough, therefore is a vaccine against Lyme borreliosis desirable.
A vaccine can contain a whole variety of different antigens. Examples of antigens are whole-killed or attenuated organisms, subfractions of these organisms/tissues, proteins, or, in their most simple form, peptides. Antigens can also be recognized by the immune system in form of glycosylated proteins or peptides and may also be or contain polysaccharides or lipids. Short peptides can be used since for example cytotoxic T-cells (CTL) recognize antigens in form of short usually 8-11 amino acids long peptides in conjunction with major to histocompatibility complex (MHC). B-cells can recognize linear epitopes as short as 4-5 amino acids, as well as three-dimensional structures (conformational epitopes). In order to obtain sustained, antigen-specific immune responses, adjuvants need to trigger immune cascades that involve all cells of the immune system. Primarily, adjuvants are acting, but are not restricted in their mode of action, on so-called antigen presenting cells (APCs). These cells usually first encounter the antigen(s) followed by presentation of processed or unmodified antigen to immune effector cells. Intermediate cell types may also be involved. Only effector cells with the appropriate specificity are activated in a productive immune response. The adjuvant may also locally retain antigens and co-injected other factors. In addition the adjuvant may act as a chemoattractant for other immune cells or may act locally and/or systemically as a stimulating agent for the immune system.